The mechanisms by which neurons maintain redox homeostasis, disruption of which is linked to disease, are not well known. Hydrogen peroxide, a major cellular oxidant and neuromodulator, can promote reversible oxidations of protein thiols but the scope, targets, and significance of such oxidations occurring in neurons, especially in vivo, are uncertain. Using redox phenylarsine oxide (PAO)-affinity chromatography, which exploits the high-affinity of trivalent arsenicals for protein dithiols, this study investigated the occurrence of reducible and, therefore, potentially regulatory, protein disulfide bonds in Triton X-100-soluble protein fractions from isolated nerve-endings (synaptosomes) prepared from rat brains. Postmortem oxidations of protein thiols were limited by rapidly freezing the brains following euthanasia and, later, homogenizing them in the presence of the N-ethylmaleimide to trap reduced thiols. The reducible disulfide proteome comprised 5.4% of the total synaptosomal protein applied to the immobilized PAO columns and was overrepresented by pathways underlying ATP synaptic supply and demand including synaptic vesicle trafficking. The alpha subunits of plasma membrane Na+, K+-ATPase and the mitochondrial ATP synthase were particularly abundant proteins of the disulfide proteome and were enriched in this fraction by 3.5- and 6.7-fold, respectively. An adaptation of the commonly used "biotin-switch" method provided additional support for selective oxidation of thiols on the alpha subunit of the ATP synthase. We propose that reversible oxidations of protein thiols may underlie a coordinated metabolic response to hydrogen peroxide, serving to both control redox signaling and protect neurons from oxidant stress.

中文翻译：

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突触小体的可还原二硫键蛋白质组支持蛋白质硫醇可逆氧化在维持神经元氧化还原稳态中的作用。

神经元维持氧化还原稳态的机制尚不清楚，该机制的破坏与疾病有关。过氧化氢是一种主要的细胞氧化剂和神经调节剂，可以促进蛋白质硫醇的可逆氧化，但是这种氧化的范围，目标和意义在神经元中，特别是在体内，尚不确定。使用氧化还原苯砷氧化物（PAO）亲和层析，该亲和层析利用三价砷对蛋白质二硫醇的高亲和力，本研究调查了Triton X-100可溶性蛋白质馏分中可还原的，因此具有潜在调控作用的蛋白质二硫键的存在从大鼠大脑中分离出的神经末梢（突触体）中提取。安乐死后迅速冻结大脑，从而限制了蛋白质硫醇的事后氧化。在N-乙基马来酰亚胺的存在下将其均质化以捕获还原的硫醇。可还原的二硫键蛋白质组占固定化PAO色谱柱总突触体蛋白的5.4％，并被包括突触小泡运输在内的ATP突触供需途径所代表。质膜Na +，K + -ATPase和线粒体ATP合酶的α亚基是二硫蛋白组中特别丰富的蛋白质，在该部分中分别富集了3.5倍和6.7倍。常用“生物素转换”方法的改编为ATP合酶α亚基上硫醇的选择性氧化提供了额外的支持。我们建议蛋白质硫醇的可逆氧化可能是对过氧化氢的协调代谢反应的基础，